cDNAs and Antibodies for Small GTP-binding Proteins

存在具有GTP酶活性的ras p21/ras p21样小GTP结合蛋白（小G蛋白）超家族。已经报道了超过40个成员，并且所有具有20，000和41，000之间的Mrs的成员都由单个相应的多肽组成。越来越多的证据表明，小G蛋白参与细胞生长和分化的调节。在本研究项目中，我们研究了小G蛋白的C-末端结构，纯化了小G蛋白的调节蛋白，并制备了针对小G蛋白及其调节蛋白的单克隆抗体。大多数小G蛋白都具有独特的C-末端保守基序，其中至少含有一个半胱氨酸残基。我们已经发现，smg p21 B，rhoA p21和smg p25 A的C-末端系统素残基是香叶基香叶基化的，并且这些异戊烯化对于每个小G蛋白与膜结合是必需的。我们已经部分纯化并表征了rhoA p21的香叶基-香叶基转移酶 ...更多信息 从牛脑中提取的小G蛋白有两种相互转化的形式，GDP结合的非活性形式和GTP结合的活性形式。从GDP结合到GTP结合形式的转化和逆转化分别由GDP/GTP交换和GTP酶反应诱导。我们已经纯化并鉴定了几种GDP/GTP交换蛋白（GDP解离刺激物（GDS）和GDP解离抑制物（GDI））和小G蛋白的GTP酶激活蛋白（GAP）。在这些调节蛋白中，我们克隆了smg p21 GDS、smg p25 A GDI和rho GDI的cDNA，并制备了针对它们的单克隆抗体。我们发现，在造血白血病细胞分化为造血细胞后，smg p21 mRNA的水平增加。我们还发现，smg p25 A的特异性单克隆抗体主要在神经组织中检测到，并且在PC 12细胞分化为神经元样细胞后，smg p25 A mRNA水平增加。这些结果表明，smg p21和smg p25 A可分别用作造血和神经组织来源的肿瘤的潜在肿瘤标志物。关键词：ras p21，小G蛋白，异戊烯基化，GDP/GTP交换蛋白，GTP酶激活蛋白，肿瘤标志物。少

There is a superfamily of ras p21/ras p21-like small GTP-binding proteins (small G proteins) with GTPase activity. Over forty members have been reported, and all of them with Mrs between 20,000 and 41,000 are composed of a single respective polypeptide. Evidence is accumulating that small G proteins are involved in the regulation of cell growth and differentiation. In this research project, we have investigated the C-terminal structures of small G proteins, purified the regulatory proteins for small G proteins, and made monoclonal antibodies against small G proteins and their regulatory proteins. Most of small G proteins have the unique consensus C-terminal motifs containing at least one cysteine residue. We have found that the C-terminal systeine residues of smg p21B, rhoA p21, and smg p25A are geranylgeranylated, and that these prenylation are essential for each small G protein to bind to membranes. We have partially purified and characterized geranyl-geranyl transferase for rhoA p21 … More from bovine brain. Small G proteins have two interconvertible forms, GDP-bound inactive and GTP-bound active forms. The conversion from the GDP-bound to GTP-bound form and the reverse conversion are induced by GDP/GTP exchange and GTPase reactions, respectively. We have purified and characterized several GDP/GTP exchange proteins (GDP dissociation stimulator (GDS) and GDP dissociation inhibitor (GDI)) and GTPase activating Proteins (GAP) for small G proteins. Among these regulatory proteins, we have cloned the cDNAs of smg p21 GDS, smg p25A GDI, and rho GDI, and made monoclonal antibodies against them. We have found that the levels of smg p21 mRNA increase in hematopoietic leukemiae cells after their differentiation into hematopoietic cells. We have also found that smg p25A is mainly detected inneural tissues by use of its specific monoclonal antibody, and that the levels of smg p25A mRNA increase in PC12 cells after their differentiation into neuron-like cells. These results suggest that smg p21 and smg p25A can be used as potential tumor markers for tumors derived from hematopoietic and neural tissues, respectively. Key Words : ras p21, small G proteins, Prenylation, GDP/GTP exchange protein, GTPase activating protein, Tumor markers. Less